A rapid and simple method to obtain canine peripheral blood-derived macrophages.

Macrophages play an important role in a variety of situations, including pathogen elimination, inflammation, and tissue repair. However, these cells are not fully studied in dogs, in part, due to the difficulty of efficiently isolating and culturing them in vitro. In this study, we cultured canine peripheral blood mononuclear cells (PBMCs) with 10 ng/ml of phorbol 12-myristate-13-acetate (PMA) for 5 days to obtain macrophages. A high number of round-adherent cells were obtained without the addition of any cytokine. These cells showed active phagocytic activity and a cell surface antigen profile different from dendritic cells. Our method facilitates a high yield of macrophages in a short cultivation period compared with previous studies. This method might be a powerful tool to study macrophage functions in dogs.

Hypereosinophilic syndrome in two cats.

Two cats showing chronic vomiting, diarrhea and weight loss were found to have leukocytosis with marked eosinophilia. Both cats were diagnosed with hypereosinophilic syndrome by the findings of increased eosinophils and their precursors in the bone marrow, eosinophilic infiltration into multiple organs, and exclusion of other causes for eosinophilia. Although cytoreductive chemotherapy with hydroxycarbamide and prednisolone was performed, these two cats died 48 days and 91 days after the initial presentation.

Usefulness of antidepressants for improving the neuropathic pain-like state and pain-induced anxiety through actions at different brain sites.

Clinically, it is well known that chronic pain induces depression, anxiety, and a reduced quality of life. There have been many reports on the relationship between pain and emotion. We previously reported that chronic pain induced anxiety with changes in opioidergic function in the central nervous system. In this study, we evaluated the anxiolytic-like effects of several types of antidepressants under a chronic neuropathic pain-like state and searched for the brain site of action where antidepressants show anxiolytic or antinociceptive effects. Sciatic nerve-ligated mice exhibited thermal hyperalgesia and tactile allodynia from days 7 to 28 after nerve ligation. At 4 weeks after ligation, these mice showed a significant anxiety-related behavior in the light-dark test and the elevated plus-maze test. Under these conditions, repeated administration of antidepressants, including the tricyclic antidepressant (TCA) imipramine, the serotonin noradrenaline reuptake inhibitor (SNRI) milnacipran, and the selective serotonin reuptake inhibitor (SSRI) paroxetine, significantly prevented the anxiety-related behaviors induced by chronic neuropathic pain. These antidepressants also produced a significant reduction in thermal hyperalgesia and tactile allodynia. Moreover, the microinjection of paroxetine into the basolateral amygdala or cingulate cortex reduced anxiety-related behavior, and microinjection into the primary somatosensory cortex significantly attenuated thermal hyperalgesia. These findings suggest that serotonergic antidepressants are effective for treating anxiety associated with chronic neuropathic pain and may be useful for treating neuropathic pain with emotional dysfunction such as anxiety. Furthermore, SSRIs show anxiolytic and antinociceptive effects by acting on different brain regions.

Chronic pain-induced emotional dysfunction is associated with astrogliosis due to cortical delta-opioid receptor dysfunction.

It has been widely recognized that chronic pain could cause physiological changes at supraspinal levels. The delta-opioidergic system is involved in antinociception, emotionality, immune response and neuron-glia communication. In this study, we show that mice with chronic pain exhibit anxiety-like behavior and an increase of astrocytes in the cingulate cortex due to the dysfunction of cortical delta-opioid receptor systems. Using neural stem cells cultured from the mouse embryonic forebrain, astrocyte differentiation was clearly observed following long-term exposure to the selective delta-opioid receptor antagonist, naltrindole. We also found that micro-injection of either activated astrocyte or astrocyte-conditioned medium into the cingulate cortex of mice aggravated the expression of anxiety-like behavior. Our results indicate that the chronic pain process promotes astrogliosis in the cingulate cortex through the dysfunction of cortical delta-opioid receptors. This phenomenon may lead to emotional disorders including aggravated anxiety under chronic pain-like state.

Chronic pain induces anxiety with concomitant changes in opioidergic function in the amygdala.

Clinically, it has been reported that chronic pain induces depression, anxiety, and reduced quality of life. The endogenous opioid system has been implicated in nociception, anxiety, and stress. The present study was undertaken to investigate whether chronic pain could induce anxiogenic effects and changes in the opioidergic function in the amygdala in mice. We found that either injection of complete Freund's adjuvant (CFA) or neuropathic pain induced by sciatic nerve ligation produced a significant anxiogenic effect at 4 weeks after the injection or surgery. Under these conditions, the selective mu-opioid receptor agonist [D-Ala2,N-MePhe4,Gly5-ol]-enkephalin (DAMGO)- and the selective delta-opioid receptor agonist (+)-4-[(alphaR)-alpha-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide (SNC80)-stimulated [35S]GTPgammaS binding in membranes of the amygdala was significantly suppressed by CFA injection or nerve ligation. CFA injection was associated with a significant increase in the kappa-opioid receptor agonist 2-(3,4-dichlorophenyl)-N-methyl-N-[(1S)-1-phenyl-2-(1-pyrrolidinyl)ethyl]acetamide hydrochloride (ICI199,441)-stimulated [35S]GTPgammaS binding in membranes of the amygdala. The intracerebroventricular administration and microinjection of a selective mu-opioid receptor antagonist, a selective delta-opioid receptor antagonist, and the endogenous kappa-opioid receptor ligand dynorphin A caused a significant anxiogenic effect in mice. We also found that thermal hyperalgesia induced by sciatic nerve ligation was reversed at 8 weeks after surgery. In the light-dark test, the time spent in the lit compartment was not changed at 8 weeks after surgery. Collectively, the present data constitute the first evidence that chronic pain has an anxiogenic effect in mice. This phenomenon may be associated with changes in opioidergic function in the amygdala.

Correlation p53 expression and human papilloma virus deoxyribonucleic acid with clinical outcome in early uterine cervical carcinoma.

BACKGROUND: In the present study we assessed whether expression of p53 protein or HPV DNA correlates with recurrence as well as several known prognostic factors in uterine cervical carcinoma. METHODS: Forty-nine patients with FIGO stage IA-IIB who underwent hysterectomy between 1998 and 2002 were retrospectively studied. All 49 cancer tissue samples were used for immunohistochemical study. Twenty-five of 49 cases were also examined by PCR-RFLP for detection and typing of HPV DNA. RESULTS: Twenty of 49 (40.8%) specimens demonstrated nuclear staining for p53. A significant association between p53 overexpression and age, hormonal status, FIGO stage, or recurrence was observed (p=0.02, 0.01, 0.03, 0.01). However, no significant association was found between p53 overexpression and lymph node metastases, parametrium involvement, or risk of death (p=0.18, 0.06, 0.14). Nineteen of 25 (76%) were HPV DNA-positive and 6 (24%) were negative. DISCUSSION: There was no relation between HPV DNA positivity and age, FIGO stage, lymph node metastases, parametrium involvement, recurrence, or risk of death. CONCLUSION: p53 overexpression is associated with age, hormonal status, FIGO stage, and recurrence in uterine cervical carcinoma.